Culture Line Of Mammary Epithelial Cells And Fibroblasts From Wanlin White Goat In Vitro

Local livestock and poultry breed resources are an important part of biodiversity,and their protection and development directly affect the sustainable development of China’s animal husbandry.Anhui white goat has good meat production performance,high reproductive rate,strong adaptability and high comprehensive benefits of breeding.There have been no studies of this animal’s genetic resources protected at the cellular level..In this study,the cell culture technology was used to establish a cell bank of Wanlin white goat cells and to characterize their growth characteristics,in order to lay the foundation for the conservation of germplasm resources of Wanlin white goat.The main research work and results are summarized as follows:1)In vitro culture of mammary epithelial cells of Wanlin white goat showed pebble-like growth and formed island-like monolayer aggregation,while fibroblasts showed long shuttle-shaped and fibrous growth;immunofluorescence method was used to identify the cells,and mammary epithelial cells were positive for keratin 18 and β-casein,while fibroblasts were positive for waveform protein;the growth curves of both cells were "S The growth curves of both cells were "S"-shaped,consistent with the in vitro cell culture pattern.2)The survival rate of the purified mammary epithelial cells of Wanlin White goat reached over 90% before freezing and the survival rate of the sixth generation of cells after recovery,indicating that the cells can be adapted to conventional freezing;the average survival rates fibroblasts before freezing and after recovery for different generations(P3,P6,P9)were 97.63% and 93.73%,respectively,indicating that the cells can be adapted to conventional freezing.3)Chromosome karyotype analysis of mammary epithelial cells and fibroblasts,including 2N=60,the diploid proportion of mammary epithelial cells and fibroblasts was90% and 88%,respectively,indicating that the genetic material can be stably inherited in these two cells.4)Oil red O staining of mammary epithelial cells passed 17 times,many lipid droplets were seen diffused in the cytoplasm under the microscope,and the cells still had a secretory function.5)The proliferation rate of Wanlin White goat fibroblasts in 15% fetal bovine serum and10% fetal bovine serum was similar,and the more economical 10% serum can be chosen for subsequent studies related to a larger range of goat fibroblast cultures.6)Culture between different generations of mammary epithelial cells(P6,P9,P12)and fibroblasts(P2,P5,P8)in Wanlin white goats,with the increase of generations,there is a tendency to decrease cell viability,apposition rate and stability of genetic material,and cells of low generations should be selected when cloning in Wanlin white goats.7)PCR mycoplasma detection,both mammary epithelial cells and fibroblasts of Wanlin white goats were free of mycoplasma contamination.In this study,mammary epithelial cell lines and fibroblast cell lines were successfully established.The experiment was conducted to protect the genetic resources of Wanlin white goats at the cellular level,to provide experimental material for molecular breeding by studying gene functions at the cellular level,and to provide test methods and test materials for future research on the cytology of Wanlin white goats.